“STUDY OF HEARTREGENERATION IN TADPOLES

OF THE TOAD,BUFO MELANOSTICTUS”

GOVIND GUPTAM.Sc BIOTECHNOLOGY

DEPARTMENT OF LIFE SCIENCESIASE DEEMED UNIVERSITY

SARDARSHAHR

INTRODUCTION Heart is a vital organ and It has long been known that the heart

is one of the least regenerative organs in the body. Owing to the lack of organ donors and complications associated

with immune suppressive treatments ,scientists are continuously looking for new strategies to regenerate the injured heart.

Mammals, including human, form scar tissue after cardiac damage like that caused by a heart attack. This scarring permanently impairs heart function. But certain model amphibians can regrow heart tissue after injury.

Vitamin A was found to be good model to accelerate heart regenerative ability in anuran amphibians.

It was decided to explore whether anuran amphibians(frogs & toads) could regenerate heart tissue under the influence of Vitamin A

The high regeneration ability of amphibians provides a valuable model system to gain basic information on regeneration that may be transferable to human trauma and diseases that cause damage to such structures.

Transplantation technique will open new doors in the field of cardiac tissue engineering.

In the present study both parameters will be discussed i.e. heart regeneration in vivo and in transplantation set up under the influence of Vitamin A.

MATERIALS AND METHODS Young (3 toe stage) and mature (5 toe stage)

tadpoles of the toad, were employed as experimental animals.

Experiment was completed in two series. In the first series a small cut was made in

skin on anterior ventral surface in 40 tadpoles to expose the heart and then the tip of ventricle part of heart was incised.

Operated half (20) animals were reared in tap water (controls) and remaining half (20) in vitamin A solutions (5 IU/ml) for first three days and then transferred into water.

In second series of experiment-meshed heart tissue was implanted into a pit made on mid-lateral position of tail in 40 host tadpoles. Half (20) of the operated tadpoles with implants were reared in water(control) and remaining (20) were reared in vitamin A solution(5 IU/ml) for first three days and then transferred into water. Experiment was terminated on day 15 after operation.

PHOTOGRAPH SHOWING AMPUTATION LEVEL OF VENTRICLE PART OF THE HEART .

PHOTOGRAPH SHOWING IMPLANTATION OF CARDIAC TISSUE IN MID LATERAL POSITION OF TAIL OF TADPOLES.

RESULT & OBSERVATIONSSeries I

Mode of experiment Stage of experimental animals

Groups (After operation animals

reared in)

Day of preservation No. of preserved animals

Percentage of heart regeneration

SI

(Heart regeneration in vivo) Amputation of tip of the ventricle

3 toe stage young tadpoles

Control IA

in water( 1-15 days)

3

7

15

5

5

10

40%

Vit. A treated IB

3days in Vit. A, then reared in water

3

7

15

5

5

10

65%

5 toe stage mature tadpoles

Control IC

in water( 1-15 days)

3

7

15

5

5

10

30%

Vit. A treated ID

3days in Vit. A, then reared in water

3

7

15

5

5

10

55%

Series 1: Percentage of heart regeneration in situ in the tadpole of the Toad.

Mode of experiment Stage of experimental animals

Groups (After operation animals

reared in)

Day of preservation No. of preserved animals

Percentage of heart regeneration

SII

(Implantation of mashed heart tissue

in pit on the tail region)

3 toe stage young tadpoles

Control IIA

in water( 1-15 days)

3

7

15

5

5

10

30%

Vit. A treated IIB

3days in Vit. A, then reared in water

3

7

15

5

5

10

55%

5 toe stage mature tadpoles

Control IIC

in water( 1-15 days)

3

7

15

5

5

10

20%

Vit. A treated IID

3days in Vit. A, then reared in water

3

7

15

5

5

10

45%

Series II

Series II: Cardiac Tissue Regeneration from mashed heart Tissue at ectopic site in the Tadpoles of Toad.

Heart regeneration in present results clearly demonstrate an ability of toad tadpoles to heal myocardial injuries.

Vitamin A has proved to be significant chemical model to induce and accelerate heart regeneration in both modes of experiments.

In first mode of experiment. The percentage of heart regeneration in vivo in 3 toe stage tadpoles it was 65% in

Vit-A treated cases and 40% in control (water), and in 5 toe stage tadpoles percentage of heart regeneration is 55% in Vit.A treated and 30% in control (water) cases.

In second mode of experiment The percentage of heart regeneration in vivo in 3 toe stage tadpoles it was 55%in

Vit-A treated cases and 30% in control (water) but in 5 toe stage tadpoles the percentage of regeneration is 45% in Vit. A treated and 20% in control (water) cases.

The regeneration ability declines according to the animal age. The basic pattern of heart regeneration was found similar in

both Vit-A treated as well as untreated control group tadpoles. For the study of sequential events occurred during heart

regeneration, operated animals were preserved at different time intervals (Day 5,10 and d15).By day 5 ,the wound showed proper healing (Fig.3),whereas on day 10 the wound showed rapid resolution of granulation tissue and a restoration of normal myocardial architecture.

PHOTOGRAPH SHOWING COMPLETE HEART REGENERATION ON DAY 15 AFTER OPERATION.(RH = REGENERATED HEART)

PHOTOGRAPH SHOWING COMPLETE HEART REGENERATION VENTRICLE PART ON DAY 15 AFTER OPERATION.(RV= REGENERATED VENTRICLE)

PHOTOGRAPHS OF CARDIAC TISSUE IMPLANTS ON ECTOPIC SITE SHOWING DEVELOPMENT, DIFFERENTIATION AND GROWTH OF THE IMPLANTS ON DAY 15. (20×) (DEH=DEVELOPING ECTOPIC HEART)

PHOTOGRAPH IS SHOWING CARDIAC MUSCLES CELLS OF HEART.(CMC – CARDIAC MUSCLES CELLS)

On the wound site or injury site generation of new cells observed that were indistinguishable from neighboring healthy heart cells.

Promising results obtained in the second phase of experiment. The mode of experiment was ectopic transplantation of meshed cardiac tissue. The cardiac patches could survive and beat for up to 15 days after engraftment on the tail.

CONCLUSION In light of the above results obtained in the present

study ,amphibian system can make a substantial contribution to our understanding of heart regeneration

These important areas for research have the potential to provide basic information that could be used to induce and control heart repair in mammals.

The findings also suggest that all vertebrates including human, might possess this regenerative capability and that methods could be developed to tap it.

Transplantation technique opens new doors in the field of cardiac tissue engineering.

THANKS